A. Arcangeli et al., EFFECTS OF INHIBITORS OF ION-MOTIVE ATPASES ON THE PLASMA-MEMBRANE POTENTIAL OF MURINE ERYTHROLEUKEMIA-CELLS, The Journal of membrane biology, 126(2), 1992, pp. 123-136
The membrane electric effects of N,N'-dicyclohexyl-carbodiimide (DCCD)
and vanadate were studied in murine erythroleukemia cells (MELC), com
paring the patch-clamp technique and the accumulation ratio (AR(exp))
of [H-3]-tetraphenylphosphonium (TPP+). Electrophysiological measureme
nts showed that both these inhibitors produce, at micromolar concentra
tions, a 20-30 mV hyperpolarization of resting potential (DELTA-psi(p)
) of MELC, which is abolished when the electrochemical equilibrium pot
ential of K+ (E(K)) is brought close to zero. DCCD and vanadate turned
out to have distinct targets on the plasma membrane of MELC (an H+ pu
mp and the Na+,K+-ATPase, respectively). Measurements of AR(exp) showe
d that: (i) patch-clamp measurements of DELTA-psi(p) were equivalent t
o those based on AR(exp) of antimycin-pretreated cells (AR(ANT)); (ii)
DCCD produced a strong increase in AR(ANT), that was antagonized by c
arbonyl cyanide p-trifluoromethoxyphenyl-hydrazone (FCCP) and diethyls
tilbestrol (DES); (iii) vanadate determined a marked increase in AR(AN
T) that was insensitive to FCCP, but antagonized by ouabain; (iv) incu
bation in high K+ medium (HK) brought AR(ANT) to 1.0 in the controls,
but did not lower this ratio below 3.0 in the presence of DCCD or vana
date; (v) the total amount of TPP+ taken up by the cells was in any ca
se water extractable by a freezing and thawing procedure. On the whole
, our data indicate that DCCD and vanadate hyperpolarize the MELC by i
ncreasing the K+ conductance and, at the same time, enhance the TPP+ b
inding, probably by changing the electrostatic potential profile of th
e plasma membrane. These effects seem to involve functional modificati
ons of the target pumps, apparently related to the ion-occluding state
of these enzymes.